Hematopoiesis is the process by which blood cells develop and differentiate from pluripotent stem cells in the bone marrow. This process involves a complex interplay of polypeptide growth factors (cytokines) acting via membrane-bound receptors on the target cells. Cytokine action results in cellular proliferation and differentiation, with response to a particular cytokine often being lineage-specific and/or stage-specific. Development of a single cell type, such as a platelet or erythrocyte, from a stem cell may require the coordinated action of a plurality of cytokines acting in the proper sequence.
The known cytokines include the interleukins, such as IL-1, IL-2, IL-3, IL-6, IL-8, etc.; and the colony stimulating factors, such as G-CSF, M-CSF, GM-CSF, erythropoietin (EPO), etc. In general, the interleukins act as mediators of immune and inflammatory responses. The colony stimulating factors stimulate the proliferation of marrow-derived cells, activate mature leukocytes, and otherwise form an integral part of the host's response to inflammatory, infectious, and immunologic challenges.
Various cytokines have been developed as therapeutic agents. Several of the colony stimulating factors have been used in conjunction with cancer chemotherapy to speed the recovery of patients' immune systems. Interleukin-2, .alpha.-interferon and .gamma.-interferon are used in the treatment of certain cancers. EPO, which stimulates the development of erythrocytes, is used in the treatment of anemia arising from renal failure. Factors responsible for stimulation of megakaryocytopoiesis and thrombocytopoiesis resisted definitive characterization, due in part to lack of a good source, a lack of good assays, and a lack of knowledge as to the site(s) of production until recently, despite three decades of work to isolate and characterize them. The megakaryocytopoietic factor referred to in the literature as "thrombopoietin" (recently reviewed by McDonald, Exp. Hematol. 16:201-205, 1988; and McDonald, Am. J. Ped. Hematol. Oncol. 14:8-21, 1992) has now been identified and isolated (see copending U.S. patent application Ser. No. 08/252,491; Lok et al., Nature 369:565-568, 1994; and Kaushansky et al., Nature 369:568-571, 1994; all herein incorporated by reference).
Mild bleeding disorders (MBDs) associated with platelet dysfunctions are relatively common (Bachmann, Seminars in Hematology 17: 292-305, 1980), as are a number of congenital disorders of platelet function, including Bernard-Soulier syndrome (deficiency in platelet GPIb), Glanzmann's thrombasthenia (deficiency of GPIIb and GPIIIa), congenital afibrinogenemia (diminished or absent levels of fibrinogen in plasma and platelets), and gray platelet syndrome (absence of .alpha.-granules). In addition there are a number of disorders associated with platelet secretion, storage pool deficiency, abnormalities in platelet arachidonic acid pathway, deficiencies of platelet cyclooxygenase and thromboxane synthetase and defects in platelet activation (reviewed by Rao and Holmsen, Seminars in Hematology 23: 102-118, 1986). At present, the molecular basis for most of these defects is not well understood.
Anemias are deficiencies in the production of red blood cells (erythrocytes) and result in a reduction in the level of oxygen transported by blood to the tissues of the body. Hypoxia may be caused by loss of large amounts of blood through hemorrhage, destruction of red blood cells from exposure to autoantibodies, radiation or chemicals, or reduction in oxygen intake due to high altitudes or prolonged unconsciousness. When hypoxia is present in tissue, EPO production is stimulated and increases red blood cell production. EPO promotes the conversion of primitive precursor cells in the bone marrow into pro-erythrocytes which subsequently mature, synthesize hemoglobin and are released into the circulation as red blood cells. When the number of red blood cells in circulation is greater than needed for normal tissue oxygen requirements, the level of EPO in circulation is decreased.
Severe reductions in both megakaryocyte and erythrocyte levels can be associated with the treatment of various cancers with chemotherapy and radiation and diseases such as AIDS, aplastic anemia and myelodysplasias. Levels of megakaryocytes and/or erythrocytes that become too low, for example, platelet counts below 25,000 to 50,000 and hematocrits of less than 25, are likely to produce considerable morbidity and in certain circumstances these levels are life-threatening. In addition to treating the underlying disease, specific treatments include platelet transfusions for thrombocytopenia (low platelet counts) and stimulation of erythropoiesis using EPO or transfusion of red blood cells for anemia.
Recent advances in molecular biology have greatly increased our understanding of hematopoiesis, but at the same time have shown the process to be extremely complex. While many cytokines have been characterized and some have proven clinical applications, there remains a need in the art for additional agents that stimulate proliferation and differentiation of myeloid and lymphoid precursors and the production of mature blood cells. There is a particular need for agents that stimulate the development and proliferation of cells of the megakaryocytic and erythroid lineages, including platelets and red blood cells. There is a further need in the art for agents that can be used in the simultaneous treatment of cytopenias and anemias such as those caused by destruction of hematopoietic cells in bone marrow such as in the treatment of cancer with chemotherapy and radiation, and pathological conditions such as myelodysplasia, AIDS, aplastic anemia, autoimmune disease or inflammatory conditions. The present invention fulfills these needs and provides other, related advantages.